Synthesis and biological evaluation of closo-dodecaborate ibuprofen conjugate (DIC) as a new boron agent for neutron capture therapy

Dedicated to Professor Narayan Hosmane on the occasion of his 70th birthday.
https://doi.org/10.1016/j.jorganchem.2018.02.026Get rights and content

Highlights

  • A closo-dodecaborate ibuprofen conjugate (DIC) 1 was synthesized.

  • DIC accumulated in HeLa cells in a time-dependent manner.

  • DIC showed no significant binding affinity to the ibuprofen binding site of HSA.

Abstract

A closo-dodecaborate ibuprofen conjugate (DIC) 1 was synthesized via amide bond formation between ibuprofen activated with 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide (EDCI) and 4-aminoethoxyethoxy-closo-dodecaborate 3 which was synthesized via the ring opening reaction of 1,4-dioxane-closo-dodecaborate complex 2 with ammonia in aqueous solution. DIC showed no significant binding affinity to HSA site 2, which is known as an ibuprofen binding site. DIC accumulated in HeLa cells in a time-dependent manner, and the boron concentration reached 0.68 μg/106 cells 6 h after administration.

Graphical abstract

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A closo-dodecaborate ibuprofen conjugate (DIC) 1 was synthesized and evaluated for albumin binding assay and cell-uptake using HeLa cells.

Introduction

The icosahedral symmetrical cluster, closo-dodecaborate, contains twelve boron and twelve hydrogen atoms [[1], [2], [3]]. It is a double negatively charged ionic molecule with molecular formula [B12H12]2-, and its B12 core is formed through a three-center two-electron bond. The sodium form of closo-dodecaborate is water-soluble and has low toxicity [4,5]. Indeed, mercaptoundecahydrododecaborate ([B12H11SH]2-) was developed as a boron agent and used to treat brain tumors via boron neutron capture therapy (BNCT) for many years [[6], [7], [8]].

BNCT is a unique binary therapy that was developed during the last 5–6 decades. It is based on the nuclear reaction of essentially nontoxic 10B species and low-energy thermal neutrons (0.025 eV), which yield high linear energy transfer particles, 7Li nuclei and 4He (α-particles), traveling a distance equivalent to one cell diameter (approximately 5–9 μm). The energy of these particles is approximately 2.4 MeV, which is enough to kill cells. Therefore, selective and efficient boron delivery is key to successful BNCT. We previously developed maleimide-containing closo-dodecaborate (MID) that was conjugated not only with cysteine residue–SH but also with lysine residue–NH2 in proteins. We focused on serum albumins, which are essential transporter proteins for many drugs and endogenous compounds [9], using these as boron carriers for BNCT. As expected, the MID albumin conjugate (MID-AC) accumulated selectively in mouse tumors 13 h after intravenous injection via the tail vein. Tumor growth in mice injected with MID-AC was completely suppressed two weeks after thermal neutron irradiation [10]. We also developed albumin-closo-dodecaborate conjugates by Ru(bpy)3-photocatalyzed modification of tyrosine residue [11], enabling the preparation of highly boronated albumins by conjugation through cysteine, lysine, and tyrosine residues [12]. It is known that human serum albumin has two primary drug binding sites (i.e., sites 1 and 2) located in subdomains IIA and IIIA [[13], [14], [15]]. Ibuprofen exhibits high affinity to human serum albumin (HSA) via binding to site 2, as shown in Fig. 1a [13], and we believe that the ibuprofen closo-dodecaborate conjugate will be a novel boron agent, binding to human serum albumin at site 2 for delivery to tumors. In this paper, we report the design and synthesis of closo-dodecaborate ibuprofen conjugate (DIC) 1, as shown in Fig. 1b.

Section snippets

Chemistry

The synthesis of DIC is shown in Scheme 1. According to the protocol developed by Bregadze et al. [16], the bis-tetrabutylammonium (TBA) salt of closo-dodecaborate was treated with NaBF4 in 1,4-dioxane under acidic conditions at 100 °C for 12 h, and the resulting crude product was easily purified by recrystallization from ethanol to provide 1,4-dioxane-closo-dodecaborate complex 2 in 87% yield. The ring opening of 2 proceeded in aqueous ammonia solution at 50 °C for 11 h to afford a white solid

Conclusions

We successfully synthesized closo-dodecaborate ibuprofen conjugate (DIC) 1 via ring opening of 1,4-dioxane-closo-dodecaborate complex 2 with aqueous ammonia based on Bregadze's protocol followed by EDCI-induced amide bond formation. Although DIC showed no significant binding affinity to HSA site 2, which is known as an ibuprofen binding site, it accumulated in HeLa cells in a time-dependent manner. Furthermore, the boron concentration of DIC in the cells 6 h after administration was higher than

General

NMR spectra were recorded on a Bruker Biospin AVANCE-400 (400 MHz for 1H) or AVANCE-500 (500 MHz for 1H, 125 MHz for 13C, 160 MHz for 11B) instrument in the indicated solvent; 11B-NMR spectra were obtained by a11Bsingle bond1H decoupling method. Chemical shifts are reported in parts per million (ppm) relative to the signal (0.00 ppm) of tetramethylsilane as an internal standard in CDCl3 (7.26 ppm for 1H, 77.0 ppm for 13C) or CD3CN (1.94 ppm for 1H, 118.26 ppm for 13C). Multiplicities are reported using

Acknowledgements

This work was supported by a Grant-in-Aid for Scientific Research (B) (No. 17H02202) from the Ministry of Education, Culture, Sports, Science and Technology, Japan.

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